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An intercomparison study of ELISAs for the detection of porcine reproductive and respiratory syndrome virus - evaluating six conditionally dependent tests.

ABSTRACT: Latent class analysis is a widely used statistical method for evaluating diagnostic tests without any gold standard. It requires the results of at least two tests applied to the same individuals. Based on the resulting response patterns, the method estimates the test accuracy and the unknown disease status for all individuals in the sample. An important assumption is the conditional independence of the tests. If tests with the same biological principle are used, the assumption is not fulfilled, which may lead to biased results. In a recent publication, we developed a method that considers the dependencies in the latent class model and estimates all parameters using frequentist methods. Here, we evaluate the practicability of the method by applying it to the results of six ELISA tests for antibodies against the porcine reproductive and respiratory syndrome (PRRS) virus in pigs that generally follow the same biological principle. First, we present different methods of identifying suitable starting values for the algorithm and apply these to the dataset and a vaccinated subgroup. We present the calculated values of the test accuracies, the estimated proportion of antibody-positive animals and the dependency structure for both datasets. Different starting values led to matching results for the entire dataset. For the vaccinated subgroup, the results were more dependent on the selected starting values. All six ELISA tests are well suited to detect antibodies against PRRS virus, whereas none of the tests had the best values for sensitivity and specificity simultaneously. The results thus show that the method used is able to determine the parameter values of conditionally dependent tests with suitable starting values. The choice of test should be based on the general fit-for-purpose concept and the population under study.

SUBMITTER: Schoneberg C

PROVIDER: S-EPMC8789123 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundThe aim of the study was to evaluate the performance of different newly developed and/or commercially available ELISAs for detection of PRRSV specific antibodies. Consequently, ten PRRSV negative piglets (group V) were vaccinated with a PRRSV type 2 vaccine. Blood samples were taken before as well as seven, 21 and 42 days after vaccination. At day 42 after vaccination (day 0 of the study) all of the piglets from group V and 10 non-prevaccinated PRRSV negative piglets (group N) were challenged with an HP PRRSV type 2 field strain. Blood samples were taken before and at days 3, 7, 10, 14, 21 and 28 after challenge. The success of vaccination and challenge was measured with RT qPCR. All serum samples were tested with six ELISAs for detection of PRRSV antibodies. Three of them are nucleocapsid-based, two use a glycoprotein extract and one uses inactivated whole virus as antigen. The specificity of the ELISAs was evaluated using 301 serum samples of piglets from PRRSV negative herds.ResultsThe piglets from group V tested positive by RT qPCR at day 7 after vaccination and all piglets tested positive at day 3 after challenge. PRRSV specific antibodies were seen with all nucleocapsid-based ELISAs from day 21 after vaccination onwards in group V and from day 10 after challenge in group N. The glycoprotein-based ELISAs detected antibodies from day 42 after vaccination (group V) and day 21 after challenge (group N). The agreement according to kappa-coefficient was almost perfect. The glycoprotein-based ELISAs were able to distinguish PRRSV type 2, although with some cross reactions. Regarding specificity, the ELISAs performed differently (specificity between 97.4 % and 100 %), whereas most of the ELISAs with higher sensitivity had a slightly lower specificity.ConclusionsAll tested ELISA were able to detect PRRSV antibodies in the serum of pigs vaccinated with a PRRSV type 2 vaccine and after challenge with an HP PRRSV type 2 field strain. The onset on antibody detection differed, depending on the type of antigen used in the ELISAs. Most of the ELISAs with a higher sensitivity had a lower specificity.

Project description:BackgroundIn this study, six enzyme-linked immunosorbent assays (ELISA), intended for routine porcine reproductive and respiratory syndrome virus (PRRSV) herd monitoring, are tested for their ability to detect PRRSV specific antibodies in the serum of pigs after vaccination with an inactivated PRRSV type 1 vaccine and subsequent infection with a highly pathogenic (HP) PRRSV field strain. For this reason, ten piglets (group V) from a PRRSV negative herd were vaccinated twice at the age of 2 and 4 weeks with an inactivated PRRSV vaccine. Ten additional piglets (group N) from the same herd remained unvaccinated. Three weeks after second vaccination, each of the piglets received an intradermal application of an HP PRRSV field strain. Serum samples were taken before first vaccination as well as before and 3, 7, 10 and 14 days after HP PRRSV application. All serum samples were tested for PRRSV RNA by reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) as well as for PRRSV antibodies with all six study ELISAs.ResultsAt the beginning of the study (before vaccination), all of the piglets were PRRSV antibody negative with all study ELISAs. They also tested negative for PRRSV RNA measured by RT-qPCR. From day 3 after HP PRRSV application until the end of the study, a viremia was detected by RT-qPCR in all of the piglets. On day 0 (day of HP PRRSV application), nine out of ten piglets of the pre-vaccinated group tested PRRSV antibody positive with one of the tested ELISAs, although with lower S/P values than after infection. On day 10 after HP PRRSV application, all study ELISAs except one had significantly higher S/P or OD values, respectively more positive samples, in group V than in group N.ConclusionsOnly one of the tested ELISAs was able to detect reliably PRRSV antibodies in pigs vaccinated with an inactivated PRRSV vaccine. With most of the tested ELISAs, higher S/P values respectively more positive samples after PRRSV infection were seen in the pre-vaccinated group than in the non-vaccinated.

Project description:BackgroundPorcine reproductive and respiratory syndrome virus (PRRSV) is recognized worldwide as an important and economically devastating pathogen in pig production. Although PRRSV is widespread in domestic swine, there is a lack of information regarding PRRSV infection in European wild boars (Sus scrofa). Currently available information does not provide conclusive evidence that wild boars are a reservoir of PRRSV. Nevertheless, wild boars may be likely to become infected by domestic swine through occasional direct or indirect contact. Furthermore, wild boars can act as a reservoir for infectious diseases of domestic pigs. Therefore, the objectives of the present study were to determine the virus prevalence and further explore the epidemiology and diversity of PRRSV strains present in Lithuanian wild boars over a 5-year period. A total of 1597 tissue and serum samples from wild boars inhabiting 44 districts and ten counties in Lithuania were analysed using conventional nested reverse transcription polymerase chain reaction (RT-PCR) and real-time Taqman RT-PCR for the detection of PRRSV-specific open reading frame (ORF) 1 and 6 sequences.ResultsPRRSV was highly prevalent in Lithuanian wild boar populations, with an average rate of 18.66 % using conventional RT-PCR and 19.54 % using real-time RT-PCR. PRRSV was detected in 36.71 and 41.77 % of 237 hunting grounds tested by conventional RT-nPCR and real-time RT-PCR, respectively. No statistically significant differences in PRRSV prevalence were observed by geographic area in the ten Lithuanian counties. Animals infected with PRRSV were identified in all age groups; however, significantly higher prevalence rates were identified in subadult and adult wild boars than in juveniles up to 12 months old. No positive results were obtained using conventional PCR with Type 2 specific primers. Phylogenetic analysis of the partial ORF5 region revealed that ten wild boars harboured virus sequences belonging to genetic subtypes 3 and 4 and may therefore pose a serious threat to Lithuanian pig farms in which only subtype two strains are circulating.ConclusionsThe results of virus prevalence and phylogenetic analyses strongly support the role of wild boars as a possible natural reservoir for PRRSV in Lithuania.

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An intercomparison study of ELISAs for the detection of porcine reproductive and respiratory syndrome virus - evaluating six conditionally dependent tests.

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